Chunk #41 — EXPERIMENTAL PROCEDURES — Western Blots and Antibodies

Source

IRE1α induces thioredoxin-interacting protein to activate the NLRP3 inflammasome and promote programmed cell death under irremediable ER stress.

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For protein analysis, cells were lyzed in 1x M-PER buffer (#78501, Pierce) plus 10U protease inhibitor (#P840 from Sigma) and 250 μM sodium fluoride (#S299-100, Fisher Scientific). The protein concentration of samples was determined using a Thermo BCA Assay. Western blots were performed using the Invitrogen XCell SureLock® Mini-Cell and XCell II™ Blot Module (#EI0002) plus NuPage 10% (#NP0315BOX) and 12% (#NP0341BOX) Bis-Tris precast gels. Gels were run using MES buffer (#NP0002) and transferred onto Immobilin-P transfer membrane (IPVH07850 from Millipore) using a XCell II™ Blot Module (#EI9051). See Supp methods for details on antibodies, dilutions, and detection.